This invention relates to a microinjection apparatus of a micromanipulator for injecting liquids, and to a method of controlling microinjection performed by this apparatus.
In the realm of biotechnology, there is a field devoted to the artificial manipulation of genes, cells and the like to produce a body of new genetic information which is then utilized or studied. The subjects of study in this field are multifarious and include genes, cells, nuclei, fertilized embryos, tissue and protozoa. Micro-manipulation is available as a means of physically and mechanically manipulating those of these items that can be observed by an optical microscope. For example, micromanipulation is essential when injecting a foreign gene into a plant or animal cell or nucleus and analyzing the character manifesting mechanism of the gene, when segmenting an early embryo or a morula, implanting it in an adoptive parent and forming monozygotic multiples or when transplanting another individual into the cytoplasm of a fertilized egg to form a clone.
FIG. 1 is a diagram showing the overall arrangement of a micromanipulation system according to the prior art.
As shown in FIG. 1, the system includes a base 1, a microscope 2 disposed on the base 1, a position detector 3, a fine-movement unit 4, a coarse-movement unit 5, a television camera 6, a microinjector 7, a left control box 8, a right control box 9, a camera control unit 10, a video monitor 11 and a main control unit 12.
As illustrated in the Figure, the two control boxes 8, 9 are for manipulating the left and right fine-movement and coarse-movement units 4, 5 and for controlling various functions such as measurement of the amount of an injected liquid. The microscope 2 is provided with the TV camera 6 so that the state of a cell and the micromanipulation thereof can be observed by being projected on the video monitor 11.
The arrangement of a conventional micromanipulation apparatus is as shown in FIG. 2. A miniature instrument 13 (e.g., a microneedle, micropipette or microelectrode) at the tip of the fine-movement unit is supported on a support rod 14, and a tube 15 is connected to the support rod 14. In order to make an injection into or a withdrawal from a cell, a plunger 17 of a microsyringe 16 is fed on threads by turning a knob 18, whereby positive or negative pressure can be applied.
This conventional microinjection apparatus will now be described in greater detail.
Many substances do not allow free passage through their cell membrane. Consequently, in order to examine the direct action of a substance on the protoplasm or organelle in the interior of a cell, microinjection in which the substance is injected into the cell through a capillary tube (micropipette) is the most direct method.
Though microinjection generally refers to an operation for injecting a substance into a cell, the same method can be utilized in a case where a liquid is made to act upon a specific region of a cell or tissue from the outside, or in a case where a portion of the protoplasm is sucked out of the interior of a cell or a minute object such as a small cell is sucked out. Methods of microinjection include a pressure method, in which a liquid placed inside a micropipette in advance is ejected from the micropipette by pressure, or a microelectrophoretic method, in which an electric current is passed through a micropipette and a substance within the micropipette is made to flow out along with the current.
Microinjection by way of the pressure method mentioned above will now be described.
As shown in FIG. 3, a microinjection apparatus relying upon the pressure method includes a micropipette (a miniature instrument) 21 attached to a hollow support rod 22, and a microsyringe 24 to which the support rod 22 is connected by a tube 23. The microsyringe 24 has the structure of a threaded-type syringe and is so adapted that positive or negative pressure can be applied by manually operating a knob 25.
Though different from a microinjection apparatus, an arrangement which can replace the above-described conventional micromanipulator has already been proposed in a U.S. patent application (U.S.S.N. 07/410,755) entitled "MICROMANIPULATOR" to which an invention entitled "APPARATUS FOR EFFECTING FINE MOVEMENT BY IMPACT FORCE PRODUCED BY PIEZOELECTRIC OR ELECTROSTRICTIVE ELEMENT" filed by the applicant of this application has been applied, wherein an inertial member is accelerated by applying a voltage to a piezoelectric/electrostrictive element and the reaction is utilized to cause movement of a miniature instrument, thereby making possible fine movement on the order of 10 nm-10 .mu.m in a positive or negative direction. However, in the conventional microinjector described above, control of the microinjector depends upon the manual operation of a knob. As a result, the microinjector is difficult to operate and is unsatisfactory for accurate injection. In other words, in order to perform accurate injection, proficiency in the characteristics of the microinjector is required and a high level of skill is required also for the operating technique.
In addition, fine resolution cannot be obtained by an injection operation using a manually operated knob.
Furthermore, in case of the above-described microinjector, the tube connecting the microsyringe and the support rod is of considerable length. This requires a corresponding amount of space and slows down response by a commensurate amount.